parent handbook (pdf) - st. charles community college

EEELLLIIISSSAAA---TTTEEEKKK®®®

CCOOOOKKEEDD MMEEAATT SSPPEECCIIAATTIIOONN KKIITTSS

For the Qualitative Detection of Animal Species Content in Cooked and Canned Meat & Poultry Products

by Enzyme-Linked ImmunoSorbent Assay (ELISA)

INSTRUCTIONS FOR USE

ELISA-TEK® Cooked Beef Kit - Catalog # 510611 ELISA-TEK® Cooked Pork Kit - Catalog # 510621

ELISA-TEK® Cooked Poultry Kit - Catalog # 510631 ELISA-TEK® Cooked Sheep Kit - Catalog # 510641 ELISA-TEK® Cooked Horse Kit - Catalog # 510651 ELISA-TEK® Cooked Deer Kit - Catalog # 510661

ELISA-TEK® Cooked 3 Species Kit - Catalog # 510603 ELISA-TEK® Cooked 4 Species Kit - Catalog # 510604

ELISA-TEK® Cooked Custom Kit - Catalog # 510601



1 ELISA Technologies, Inc.

2501 NW 66th Court, Gainesville, FL 32653 USA Tel: (352) 337-3929 Fax: (352) 337-3928

CONTENTS

Page Number

2 INTRODUCTION

2 PRINCIPLE OF THE TEST

3 SAFETY/COSHH NOTE

3 KIT COMPONENTS

4 SHELF LIFE

4 KIT STORAGE AND EXTRACTED SAMPLE STORAGE

4 MATERIALS REQUIRED BUT NOT PROVIDED

5 PROCEDURAL NOTES AND PRECAUTIONS

6 SAMPLE PREPARATION AND EXTRACTION

7 PREPARATION OF A PLATE PLAN

7 PREPARATION OF KIT MATERIALS

8 PROCEDURE SUMMARY FLOW CHART

9 DETAILED ELISA PROCEDURE

11 DETERMINATION OF RESULTS

12 PERFORMANCE CHARACTERISTICS

12 SPECIFICITY

13 CROSS REACTIVITY TABLE

14 DISCLAIMER

14 REFERENCES

15 REVISION HISTORY





INTRODUCTION

Preventing adulteration of animal products with less desirable or objectionable species is important for economic, regulatory, health, and cultural reasons. The identification of meat species is performed in many countries to assure consumers that the meat and poultry they purchase is safe, wholesome, unadulterated and properly labeled, and may be of importance in various communities where the consumption of a particular meat is proscribed. The Cooked Meat Species ELISA, developed and utilized by the USDA, is based on

antibodies raised to heat-resistant, species-specific, muscle-related glycoproteins and employs the principle of the Enzyme-Linked ImmunoSorbent Assay (ELISA). ELISA-TEK® COOKED MEAT SPECIATION KITS are sensitive and specific tests designed to determine the species content in cooked meat and poultry products. The ELISA-TEK® COOKED MEAT SPECIATION KITS have been formatted and refined for ease of use and will meet or exceed USDA-FSIS protocol standards for the Cooked Meat Species ELISA. This kit may also be used with the ELISA-TEK© protocol which allows for flexibility in sample matrix and slightly higher sensitivity. NOTE: This manual contains information from "Detection of poultry and pork in cooked and canned meat foods by enzyme-linked immunosorbent assays" by Ronald G. Berger1 as found in the United States Department of Agriculture Food Safety and Inspection Service Microbiology Laboratory Guidebook2. Following the USDA Procedures given in this manual and the USDA-FSIS MLG2 will enable the user of the ELISA-TEK® Cooked Meat Speciation Kit to produce assay results in accordance with official USDA-FSIS protocols.

PRINCIPLE OF THE TEST

ELISA-TEK® COOKED MEAT SPECIATION KITS will identify the species of animal tissue

used as ingredients in cooked meat foods by employing a simple extraction of samples and detection by sandwich-type Enzyme-Linked ImmunoSorbent Assay.

With this type of ELISA, a capture antibody is bound to the plastic of the microplate wells. The sample extract is added to the wells and tissue antigens of the species of interest are immobilized by the capture antibody in the wells. After washing to remove unbound material, a secondary antibody with the same species specificity as the capture antibody is added. This biotinylated antibody binds to any antigens captured on the plate; excess unbound biotinylated antibody is removed by washing. Streptavidin-horseradish peroxidase conjugate is then added to the wells and binds to the immobilized biotinylated antibodies. Unbound conjugate is removed by washing. A substrate (ABTS) for the enzyme is then added to the wells. If species antigens are present in the test sample, a green color will develop as a result of the action of the bound enzyme on the substrate. The color development may be read photometrically by using a microplate reader. NOTE: The Cooked Meat Species ELISA is intended for use as a qualitative test only. The color development is proportional to the original amount of species antigen in the extract, but one should not attempt to quantify the amount of species tissue in a sample based on this assay. Variations in sample content (e.g., % lean tissue, % moisture, % fat, etc.) and sample treatment (e.g., cooking times, temperatures, etc.) will affect the amount of detectable antigen in the extract.





SAFETY/COSHH NOTE The techniques of “Good Laboratory Practice” should be employed when using this kit; if such practices are used, the reagents constitute a very low potential risk to health. Safety clothing (lab coat, glasses and gloves if necessary) should be worn and skin contact with reagents avoided; do not ingest. Any contact with skin/eyes should be treated by washing/irrigation. It is also important to be aware of the allergic, toxic or infectious potential of analytical samples.

KIT COMPONENTS

NOTE: These kits are sold in a range of test combinations containing one or more sets of species-specific reagents. Exact kit contents will therefore vary depending on the type of kit received.

A. ONE ANTIBODY-COATED MICROWELL MODULE comprised of twelve single column

strips of eight microwells each (96 test wells total), held in a plastic frame and packed in a laminate pouch with desiccants. The interior of each microwell has been coated with a calibrated amount of species-specific antibody, dried, and labeled according to its specificity.

B. THREE or MORE vials of SPECIES CONTROL containing 1.5 mL each of a species control

in buffered solution with 0.04% sodium azide as a preservative. Each serves as a positive control in the appropriate test, and can also be used as a negative control in any other Cooked Meat Species test.

C. THREE or MORE vials of BIOTINYLATED ANTI-SPECIES ANTIBODY containing 1.0 mL of

a calibrated, buffered, antibody solution containing carrier serum, a wetting agent, and 0.04% sodium azide as a preservative.

D. ONE vial of STREPTAVIDIN-PEROXIDASE CONJUGATE containing 6.0 mL of conjugate in

a buffered solution. E. ONE vial of ABTS CONCENTRATE containing 1.1 mL of a twenty-five-fold (25×) ABTS

solution. F. ONE vial of PEROXIDE CITRATE BUFFER containing 12.0 mL of citrate buffer solution with

hydrogen peroxide. G. ONE vial of STOP SOLUTION containing 6.0 mL of 1.5% w/v sodium fluoride in deionized

water. H. ONE bottle of WASH SOLUTION CONCENTRATE containing 100 mL of a ten-fold (10×)

concentrate of Tris-buffered saline with a wetting agent. I. ONE INSTRUCTIONS FOR USE manual, with one BLANK WORKSHEET and RESULTS

FORM.





SHELF LIFE The shelf life of the unopened kit is indicated on the outside label. Individual component shelf lives may vary as indicated on the respective labels. Exposure of the kit and kit components to ambient or elevated temperatures (>8°C) should be minimized.

KIT STORAGE INSTRUCTIONS

ELISA-TEK® COOKED MEAT SPECIATION KITS should be stored at 2-8°C (refrigerated). DO NOT FREEZE. Kit components should be removed from refrigeration and brought to room temperature (~20-25°C) before beginning the assay. Return unused components to refrigeration (2-8°C) after use. The ANTIBODY COATED MICROWELL MODULE must be kept DRY and WELL SEALED.

If the desiccant packet turns pink, it can be dehydrated by placing in a 100°C oven until the desiccant changes to dark blue in color. Alternately, the desiccant can be replaced or the microwell module may be stored in a desiccation chamber at 2-8°C (refrigerated).

EXTRACTED SAMPLE STORAGE Sample Extracts may be stored at 2-8°C for up to 36 hours. If prolonged storage is required, the extracts must be kept frozen. They will remain stable for several months when stored at -20°C.

MATERIALS REQUIRED BUT NOT PROVIDED

Reagents: Purified water, Sodium chloride

Equipment: Miscellaneous laboratory plastic and/or glassware, equipment for sample

handling/extraction and containers suitable for meat extracts; Whatman no. 4 or similar filter paper for clarifying sample extract; Precision micropipette and tips capable of delivering 25, 50 and 100 microliter volumes; Wash bottle for washing microwells between reagent additions. NOTE: USDA Procedures specify a dual wavelength microwell plate reader, fitted with 414 nm (405-420 nm acceptable) and 492 nm (485-500 nm acceptable) filters.

Optional: Domestic blender, mincer or stomacher and suitable sample preparation

bags; Centrifuge and appropriate centrifuge tubes; Precision repeating pipette, and tips capable of delivering 25, 50 and 100 microliter volumes; Precision multichannel pipette and tips capable of delivering 25, 50 and 100 microliter volumes; Microwell washer.





PROCEDURAL NOTES AND PRECAUTIONS

1. Review the complete Instructions for Use before performing the Cooked Meat Species

ELISA. 2. ELISA-TEK® COOKED MEAT SPECIATION KITS are intended to be used as an integral

unit. The components have been calibrated and optimized to produce consistent results. Components from other kits and/or lots should not be interchanged as they may alter the precision of the assay.

3. Each microwell strip may be used only once. 4. It is not necessary to perform the immunoassay under sterile conditions. 5. All components and test specimens should be at ambient temperature (~20-25°C) before

testing begins. 6. Mix all reagents and test specimens thoroughly before use by gentle repeated inversions or

swirling. DO NOT SHAKE. 7. Once testing has started, all steps should be completed without interruption. 8. Care must be taken not to cross-contaminate wells. A new pipette tip must be used for

each sample and control. Do not touch the top of the wells with your fingers or pipette tips. 9. Do NOT allow the conjugate to mix with the substrate. If plastic troughs are used to

disperse conjugate and substrate solutions ensure that they are always kept separate. 10. The knife, cutting surface, and hands must be thoroughly cleaned and rinsed between

samples and controls to avoid cross-contamination. 11. Incomplete well washing will adversely affect the outcome. 12. All samples to be tested must be cooked. If uncooked samples are to be tested, or if it is

suspected that the sample is not fully cooked, it is advisable to heat the extract (meat/saline mixture) in a water bath at 95-100°C for 15 minutes prior to mixing and filtration/centrifugation.

13. It is advisable to number each strip/column with a pencil on the upper frosted edge of the

strip. This preserves the identity of the strips should they become detached from the frame.





SAMPLE PREPARATION AND EXTRACTION

Prepare a saline solution (0.9% w/v sodium chloride in deionized water, e.g., 9 g NaCl in 1L H2O) for use in the extraction of meat samples.

NOTE: In view of the sensitivity of the method, care must be taken at this stage to prevent cross-contamination of samples. Any equipment, utensils, containers, or surfaces used must be thoroughly washed or discarded between extractions. NOTE: If using the ELISA-TEK protocol, it is recommended that the end user prepare a negative control which is matrix-matched to the unknown sample(s). This negative control is prepared in the same way as the unknown sample(s).

1. Prepare a product for testing by dicing or finely chopping.

2. Weigh out 5 grams of the diced sample into a clean stomacher bag or whirl-pak bag, tube,

or beaker.

3. Add 10 milliliters of the prepared saline solution (0.9% w/v NaCl).

4. Place bag and contents into a stomacher for 60 seconds. Alternatively, the mixture may be kneaded or manually homogenized.

5. Remove from the stomacher and leave undisturbed for 1 hour at room temperature.

NOTE: All samples to be tested must be COOKED. If uncooked samples are to be tested, or if it is suspected that the sample is not fully cooked, it is advisable to heat the extract (meat/saline mix) in a water bath at 95-100°C for 15 minutes prior to mixing and centrifugation/filtration.

NOTE: Depending on the type of sample, a clear liquid may appear above the settled (meat) layer; alternatively, a thin slurry may be obtained. If necessary, clarify the extract solution by filtration or centrifugation, as described in step 6.

6. Filter a portion of the liquid through a Whatman 4 (or similar) filter paper. Alternatively, you

may pour off some of the liquid into a centrifuge tube and centrifuge at 10,000 ×g for 10 minutes.

7. The clear supernatant tissue extract is now ready to be used in the ELISA.

NOTE: If the sample has a high fat content, the clear supernatant above the settled meat layer may be beneath a layer of fat. Avoid transferring the fat. It may be appropriate to carefully remove a portion of the aqueous solution using a clean (e.g. Pasteur-type) pipette into a second, clean container prior to analysis.





PREPARING A PLATE PLAN

Each ELISA-TEK® COOKED MEAT SPECIATION KIT can be used as a 96-well unit or may

be divided into a variety of strip formats depending on the number of samples to be analyzed, the species to be tested, and the number of replicates desired. IT IS IMPORTANT to prepare a

test layout showing the wells you will use for controls and samples in the following protocol. Ideally each assay will include replicates of a 100% POSITIVE TISSUE CONTROL, a 1% POSITIVE TISSUE CONTROL, and one or more NEGATIVE TISSUE CONTROLS. For the POSITIVE CONTROL, use the appropriate kit control (e.g., pork in the pork test) or a lean tissue extract of the species of interest. For the 1% POSITIVE TISSUE CONTROL, use a 1% dilution (e.g., 10 µL control + 990 µL normal saline) of the appropriate species positive control or of a lean tissue extract of the appropriate species. For the NEGATIVE CONTROLS, use one or more appropriate kit controls (e.g., beef in the pork test) and the matrix-matched negative control provided and prepared by the end user. Decide on the number of replicates of each control and sample extract you will run. For screening samples, single or duplicate microwells for each control and sample extract may be adequate.

NOTE: USDA-FSIS protocols require use of quadruplicate microwells for each control and for sample extracts that are potential violations.

PREPARATION OF KIT MATERIALS

A. ANTIBODY COATED MICROWELL MODULE: Open the foil pouch. Referring to your

plate plan, select the desired number of strips for each named species and fit into a spare frame. Replace the remaining frame and strips in the pouch, taking care that the desiccant is present, and reseal the pouch carefully. B. CONTROLS: Ready to use as 100% controls; dilute appropriate control to prepare 1%

positive control (e.g., 10 µL of control into 990 µL of prepared saline). C. ANTI-SPECIES BIOTINYLATES: Ready to use. D. STREPTAVIDIN-PEROXIDASE CONJUGATE: Ready to use. E/F. ABTS CONCENTRATE and PEROXIDE CITRATE BUFFER: Mix contents of each

separate vial by inversion. DO NOT SHAKE. ABTS is supplied as a 25× concentrate and must be diluted in PEROXIDE CITRATE BUFFER to prepare WORKING ABTS SOLUTION.

-For 96 test wells add 0.5 mL (500 µL) of ABTS CONCENTRATE to the 12.0 mL of PEROXIDE CITRATE BUFFER. Stopper the vial and mix well by gentle, repeated inversion or swirling.

-For any other number of test wells, dilute 25× ABTS CONCENTRATE with PEROXIDE CITRATE BUFFER (i.e., for each strip of 8 test wells, pipette 0.6 mL (600 µL) PEROXIDE CITRATE BUFFER into a clean container and add 25 µL ABTS CONCENTRATE. Mix well by swirling gently.

NOTE: Dilutions of ABTS CONCENTRATE should be made fresh just prior to use (e.g., during the

avidin-peroxidase conjugate incubation).

G. STOP SOLUTION: Ready to use. H. WASH SOLUTION CONCENTRATE: WASH SOLUTION CONCENTRATE is supplied as a 10× concentrate and requires dilution in distilled/deionized water to prepare WORKING WASH SOLUTION (e.g., 100 mL WASH SOLUTION CONCENTRATE to 900 mL distilled/deionized water).





ENZYME IMMUNOASSAY PROCEDURE: SUMMARY FLOW CHART

PROCEDURE VOLUME TIME DESCRIPTION

Addition 100 µL - Pipette NORMAL SALINE, CONTROL & SAMPLE

EXTRACTS into respective test wells

Incubate - 1 h Incubate at room temperature

Wash - - Wash each well 3 times using WORKING WASH SOLUTION

Addition 25 µL - Pipette ANTI-SPECIES BIOTINYLATE into all test wells

Incubate - 1 h Incubate at room temperature

Wash - - Wash each well 3 times using WORKING WASH

SOLUTION

Addition 25 µL - Pipette AVIDIN PEROXIDASE CONJUGATE into all test wells

Incubate - 30

min Incubate at room temperature

Wash - - Wash each well 6 times using WORKING WASH

SOLUTION

Addition 50 µL - Pipette WORKING SUBSTRATE SOLUTION into all test wells

Incubate - 30

min

5-30 min

ELISA-TEK PROTOCOL: Incubate 30 minutes at room temperature OR USDA PROTOCOL: Incubate at room temperature until 414-492 O.D. of positive control is between 0.450-0.500

Addition 50 µL - Pipette STOP SOLUTION into all test wells and mix by

gently rotating the microplate

Results - - Measure absorbance of each well at 414-492 nm using a microplate reader and perform data analysis





DETAILED ENZYME IMMUNOASSAY PROCEDURE

1. Remove your ELISA-TEK® COOKED MEAT SPECIATION KIT from the refrigerator. Remove

the reagents from the box and allow them to reach room temperature before starting the test. 2. Prepare the sample extracts (pg. 6), matrix-matched negative control sample (if using) and

other kit materials (pg. 7). 3. Remove the MICROWELL MODULE from its pouch and, referring to your plate plan, (see

page 7) select the desired number of strips for each named species and fit into a spare frame. Replace the remaining frame and strips in the pouch, taking care that the desiccant is present, and reseal the pouch carefully. Using a pencil, number the strips in sequence on the upper frosted edge; this preserves the identity of the strips should they become detached from the frame.

4. Using a precision micropipette, add 100µL of normal saline in each of the wells selected as

blanks. NOTE: Be sure to use a clean pipette tip for each control and sample to be tested. 5. Add 100 µL of each NEGATIVE CONTROL in each of the selected wells. 6. Add 100 µL of 1% POSITIVE CONTROL in each of the selected wells. 7. Add 100 µL of 100% POSITIVE CONTROL in each of the selected wells. 8. Add 100 µL of each SAMPLE EXTRACT in each of the selected wells. Avoid transferring any

fat from sample preparations to the wells. 9. Mix the plate gently by hand; cover. Allow to stand at room temperature for 1 hour. 10. At the end of the incubation period, empty the wells by flicking into the sink. Then carefully fill

all wells with WORKING WASH SOLUTION using a reagent wash bottle. Repeat the emptying and filling of each well twice more and then emptying into the sink. Invert the aspirated/emptied plate and rap sharply several times onto a soft paper towel placed on the lab bench.

NOTE: When inverting the plate, be sure to squeeze the plastic frame at the center of the long edges to prevent the strips from falling out of the frame.

Alternately, place the plate on the carrier of the microplate washer, (or individual strips for a strip washer) which has been primed with WORKING WASH SOLUTION and set to deliver 300 µL per well. Wash and aspirate the plate 3 times. Invert the aspirated/emptied plate and rap sharply several times onto a soft paper towel placed on the lab bench.





DETAILED ENZYME IMMUNOASSAY PROCEDURE Cont’d 11. Add 25 µL of ANTI-SPECIES BIOTINYLATE to each microwell of the relevant (same species)

ANTIBODY SENSITIZED MICROWELL STRIP(S). Work from the top to the bottom of each strip. Place a fresh tip in the micropipette and repeat the biotinylate additions as necessary for each species being run.

NOTE: Observe that the bottom of each well is covered with liquid. If not, gently tap the edge of the plate until this is accomplished. Avoid getting any antibody on the sides of the wells.

12. Cover the plate and leave at room temperature for 1 hour. 13. Repeat wash step 10. 14. Add 25 µL of AVIDIN PEROXIDASE CONJUGATE to each well. Again, observe that the

bottom of each well is covered with liquid and that no conjugate sticks to the sides of the wells.

15. Cover the plate and leave at room temperature for 30 minutes. 16. Repeat wash step 10, except wash six times instead of three. 17. Add 50 µL of the WORKING ABTS SOLUTION to each microwell (see also preparation of

WORKING ABTS SOLUTION as described on page 7, E/F). 18. ELISA-TEK PROTOCOL: Cover the plate and incubate at room temperature for 30 minutes

OR

USDA PROTOCOL: Observe the microwells containing the 100% POSITIVE CONTROL for visual color change. When color change is observed, place the plate on the reader carriage. Read and obtain 414-492nm O.D. values for the wells. Continue to read until 100% POSITIVE CONTROL O.D. values read in a range of 0.450 to 0.500 (refer to USDA-FSIS MLG2 for more detailed instructions).

19. Quickly add 50 µL of STOP SOLUTION to each well. 20. Mix the plate gently by hand to distribute the STOP SOLUTION and to prevent further color

development. 21. Read the completed assay with the aid of a microplate reader at O.D. 414-492.





DETERMINATION OF TEST RESULTS Program your microplate reader to read absorbance at 414nm (405-420nm acceptable) and then read and subtract the absorbance at 492nm (485-500nm acceptable) from the 414nm reading.

1. Place the microplate on the reader carriage and blank the instrument on the selected blank (normal saline) wells. Alternately, read all wells as raw O.D. values and manually subtract the average value of the normal saline wells from the average O.D. values of the controls and samples.

2. Read and obtain a printed copy of the absorbance values for each well.

3. Determine the blank-subtracted mean absorbance value and standard deviation of each of the POSITIVE CONTROL, 1% POSITIVE TISSUE CONTROL, NEGATIVE CONTROLS, and SAMPLE wells.

4. In order for the assay to be valid according to the ELISA-TEK PROTOCOL the following

conditions must be met: the blank-subtracted mean O.D. of the 1% positive control of the appropriate species must be greater than 0.250, the blank-corrected mean O.D.s of each of the negative controls must be less than 0.150, and the standard deviation of the replicates must be less than 0.060. If these conditions are met then the test is valid. Otherwise the test is invalid, and should be repeated.

5. In order for the assay to be valid according to USDA PROTOCOL the following conditions must be met: the blank-subtracted mean O.D. of the 100% POSITIVE CONTROL of the appropriate species must be greater than 0.600, the blank-corrected mean of the negative controls must be less than 0.060, and the standard deviation of the replicates must not be more than 0.060. If these conditions are met then the test is valid. Otherwise the test is invalid, and should be repeated.

6. If the assay is determined to be valid by the above parameters, the unknown samples may be classified as follows:

ELISA-TEK PROTOCOL:

There are two criteria which can be used to determine the status of unknown samples by this protocol. It is up to the end user to decide which criterion is valid for their samples. The kits have been validated to meet criterion 1 in our laboratory.

CRITERION 1 – Any sample whose mean blank-corrected O.D. value, minus three standard deviations, is greater than 0.250 will be considered positive; all other samples are negative. CRITERION 2 – Any sample whose mean blank-corrected O.D. value minus three standard deviations is higher than 0.100 and higher than the mean O.D. value of the highest negative control (either kit-supplied or the matrix-matched control prepared by the end user) plus three standard deviations is positive; all other samples are negative. It is recommended that the end user extract and test an appropriate matrix-matched negative control in order to use this criterion.

USDA PROTOCOL

Any sample whose mean blank-corrected O.D. value, minus three standard deviations, is greater than 0.250 will be considered positive; all other samples are negative. Report Below Regulatory

Action Levels (BRAL) for meat species or sample types that are known to not cross-react and that are positive according to ELISA-TEK Protocol Criterion 2 (above).





PERFORMANCE CHARACTERISTICS

ELISA-TEK® COOKED MEAT SPECIATION KITS, when used as directed, will produce results that meet USDA criteria, and will identify cooked meat and poultry samples containing the species of interest at levels of approximately 1% or greater.

In our laboratory, uncooked, lean meat samples prepared as directed gave positive responses when diluted 1:100 in negative meat extracts (i.e. an approximation of a sample containing 1% of the meat of interest).

ELISA-TEK® COOKED MEAT SPECIATION KITS are designed to give optimum performance at room temperature (~20-25 °C). Performance of the test above or below these temperatures may necessitate either a reduction or extension (respectively) of incubation times in order to achieve the desired degree of color of the 100% POSITIVE CONTROL (O.D. = 0.600 to 0.900 @ 414-492 nm).

When the kit and operator perform properly, the NEGATIVE CONTROL wells in each species test should appear virtually colorless to the naked eye while the POSITIVE CONTROL in each test will give a distinct medium green coloration.

Significant visible color (O.D. @ 414-492 nm > 0.150) in any of the blank or negative control wells may indicate contamination of the WORKING ABTS SOLUTION or splashing of the PEROXIDASE CONJUGATE during addition to adjacent wells. Such coloration of the negative control wells is an indication of a problem during the performance of the test and any results from that test should be interpreted with caution.

SPECIFICITY

Each set of species specific reagents has been tested against a panel of meat samples for cross-reaction and has been found to produce negative responses to the heterologous species samples.

It should be noted that samples of the feathered species (e.g. chicken, turkey, game hen, goose, duck etc.) all produce positive responses in the COOKED POULTRY assay. Similarly, a bison sample will produce a positive result in the COOKED BEEF assay, and a goat sample will produce a positive response in the COOKED SHEEP assay. Further details of responses to other and closely related species is available on request.





CROSS REACTIVITY TABLE

BEEF PORK POULTRY SHEEP HORSE DEER

KIT KIT KIT KIT KIT KIT

BEEF +++++ - - - - -

PORK - +++++ - - - -

CHICKEN - - +++++ - - -

SHEEP - - - +++++ - -

HORSE - - - - +++++ -

DEER* - - - - - +++++

COW'S MILK

- - - - - -

SHEEP’S MILK

- - - - - -

BUFFALO ++++ - - - - -

BOAR - +++++ - - - -

TURKEY - - +++++ - - -

GOAT - - - +++ - -

EGG** - - - - - -

*North American White Tail Deer

**Chicken Egg (albumin and yolk)

“-“ = Negative response to 100% lean skeletal muscle sample

“+++++” = Maximum Reaction

“+++” = Medium Reaction

Limit of Detection: < 1% for meat (all species)





DISCLAIMER

ELISA Technologies, Inc. ensures that its products are made from high quality raw materials but can make no warranty, expressed or implied, as to their suitability other than to qualitatively detect cooked meat species antigen content when used exactly in accordance with these instructions. Reminders are included as to the safe handling of materials and reagents, proper storage of material and reagents, as well as to use universal laboratory safety protocols and procedures. Use of the kit for any other purpose is considered outside its intended use.

Any damages, including consequential or special damage or expense arising directly or indirectly from using this product, are limited to replacement value of the kit at ELISA Technologies, Inc. discretion.

REFERENCES

1. Berger RG, Mageau RP, Schwab B and Johnston RW (1988). J. Assoc. Off. Anal. Chem. 71(2):406-409. 2. USDA-FSIS-OPHS (2/10/2005) Laboratory Guidebook MLG17.02, Revision 2.





REVISION NOTES 120119: Moved note on heating samples to appropriate order below step 5; decreased centrifugation time from 15m to 10m.

140601: Replaced old logo with new logo 140728: Replaced the word “ethnic” with the word “cultural” on page 3 141010: Replaced the picture on the front cover to represent new packaging

141106: Removed the inclusion of thimerosal as a preservative in the Wash Solution Concentrate 150622: Edited for clarification.

MANUFACTURED BY:

ELISA Technologies, Inc. 2501 NW 66th Court Gainesville, Florida 32653 USA Tel: (352) 337-3929 Fax: (352) 337-3928 [email protected] www.elisa-tek.com

rev.150622

http://www.elisa-tek.com/

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